E-Gel™ EX Agarose Gels, 2%, 10 Gels/Pk. - FAQs

I am not seeing any current when I try to run my E-Gel EX agarose gels. Why is this?

Here are some common reasons why your gel is not running properly:

- Copper contacts in the base are damaged due to improper use. Make sure the copper contacts in the base are intact.
- An expired or defective gel cassette was used.
- The E-Gel EX cassette was not inserted properly into a base.
- An incorrect adaptor was used.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

My E-Gel agarose gel has speckles when viewed in the imager. What should I do?

Here are some suggestions:

- Try cleaning the cassettes with alcohol and Kimwipes wipers.
- Try cleaning the camera lens.
- Try to adjust the exposure time and brightness options of the documentation system you are using.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

My sample is leaking from the wells when running my E-Gel agarose gels. What happened?

Please ensure that you have not overloaded the well and that the wells were not damaged during comb removal.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

I accidentally stored my E-Gel agarose gels at 4 degrees C instead of room temperature. Can I still use them?

While we recommend storage at room temperature, these gels will still be usable. Bring the gels to room temperature prior to the run for optimal conditions.

Find additional tips, troubleshooting help, and resources within ourNucleic Acid Gel Electrophoresis and Blotting Support Center.

How can I get better separation of my bands?

First check the percentage of your agarose gel. A higher percentage will help you to resolve smaller molecular weights while a lower percentage will help you to resolve larger molecular weights.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

Do you have a protocol for transferring RNA from my E-Gel EX Agarose Gels for Northern blotting?

Here is a suggested protocol:

- Prewet a nylon membrane suitable for use with RNA using 5X SSC buffer.
- Using the E-Gel Opener, remove the E-Gel Agarose Gel from the cassette.
- Soak gel 2 times for 10 minutes in 5X SSC, 10 mM NaOH at room temperature.
- Using standard techniques, assemble a capillary transfer device using 5X SSC, 10 mM NaOH as the transfer buffer.
- Transfer should be complete after 2 hours. Remove the membrane from the transfer setup.
- Rinse the membrane for 5 minutes in 5X SSC.
- Place the membrane on filter paper to dry (2-4 minutes). Bake the membrane for 30 minutes at 80°C under vacuum or fix RNA to the membrane using a UV crosslinker.
- Place the membrane between two pieces of blotting paper and seal in a hybridization bag. Store in a cool dry place.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

What loading buffer should I use for my E-Gel agarose gels?

Loading buffer is optional. Samples can be loaded directly into the wells if no buffer is used, or you can dilute them with deionized water or TE buffer. If you want to use a loading buffer, please see the recipes below:

E-Gel agarose gels (including EX)
10 mM Tris-HCl, pH 7.5
1 mM EDTA
0.005% bromophenol blue
0.005% xylene cyanol FF
E-Gel CloneWell II and E-Gel SizeSelect II agarose gels
10 mM Tris-HCl, pH 7.5
1 mM EDTA

Alternatively, you can use 10X BlueJuice Gel Loading Buffer or TrackIt Loading Buffer. Dilute this buffer 50- to 200-fold to obtain optimal results with E-Gel agarose gels.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

How much DNA can I load for my E-Gel Agarose Gels?

To determine the amount of DNA to load per well for your specific E-Gel agarose gel, please refer to the table on page 14 of the E-Gel Power Snap Electrophoresis System user guide (https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0017050_egel_powersnapsystem_UG.pdf).

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

How do I acquire and analyze data for my E-Gel agarose gels?

We offer our E-Editor Software, which can help you align images after a gel run. The E-Editor 2.0 Software is only available for PCs, but the older E-Gel 96 Editor software is still available for the Mac operating system and can align images from E-Gel 96 and E-PAGE 96 agarose gels. However, the original software is not compatible with E-Gel 48 or E-PAGE 48 agarose gels. Please go to www.thermofisher.com and enter "E-Editor software" in the main search to download the E-Editor Software. You can use the E-Gel Imager System for data analysis.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

What are some of the important differences between the low-throughput and high-throughput E-Gel agarose gel systems?

p>High-throughput E-Gel agarose gels have staggered wells, and are based on a neutral-pH internal buffer system as opposed to an ion exchange matrix. High-throughput E-Gel agarose gels cannot be opened, and should be run on the E-Gel E-Base system.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

I do not have enough samples to fill all wells in my E-Gel agarose gel. Can I leave some wells empty?

If you do not have enough samples to use all the wells, fill each empty well with the same volume of water as the loaded samples. For E-Gel CloneWell II agarose gels and E-Gel SizeSelect II agarose gels, it is important to add the water according to the respective manual.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Gel Electrophoresis and Blotting Support Center.

Do you have a protocol for RNA electrophoresis on your E-Gel agarose gels?

Our E-Gel agarose gels can be used for either DNA or RNA separation. RNA separation occurs under non-denaturing or denaturing conditions. Please note, our gels are not QC tested for the presence of RNases. See our suggestions below for running your non-denaturing or denaturing samples:

Non-denaturing conditions
1. Mix RNA sample with 15 µL of RNase-free water.
2. Do not heat. Load the entire sample onto the E-Gel agarose gel.
3. Electrophorese for 30 min.

Denaturing conditions
1. Mix 15 µL of RNA loading buffer with 1-5 µL of RNA (1-5 µg).
2. Heat samples at 65 degrees C for 10 min to denature RNA.
3. Place samples on ice immediately after heating.
4. Load entire sample onto E-Gel agarose gel.
5. Electrophorese for 30 min.

Note:
- The only denaturing agent that is compatible with the E-Gel system is formamide, 50-95%. Heating the sample for 5 min at 65 degrees C should be sufficient for denaturing. Using other denaturing agents like glyoxal, formaldehyde, or urea will result in very poor separation and band morphology on E-Gel agarose gels.
- Additionally, we do not recommend running samples with RNA loading buffer on the same gel as samples loaded with water.
- E-Gel EX Double Comb agarose gels and E-Gel Double Comb agarose gels with SYBR Safe stain have not been tested for use in RNA applications.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

What is the special feature of E-Gel EX agarose gels?

E-Gel EX agarose gels separate DNA faster, offer enhanced sensitivity, and provide added flexibility. The stain within the EX gels is proprietary (not based on SYBR technology), though it has the same spectral properties as SYBR stains. The E-Gel EX gels have a sensitivity that is 5X greater than gels using ethidium bromide.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

I would like to take the gel out of the E-Gel agarose gel cassette. How can I do this?

We recommend using the E-Gel Opener (Cat. No. G530001) for the SYBR and ethidium bromide E-Gel agarose gels. The E-Gel EX agarose gels, E-Gel SizeSelect agarose gels, and E-Gel GO! agarose gels can be opened with the Gel Knife (Cat. No. EI9010). We do not recommend opening the E-Gel 48 agarose gels or the E-Gel 96 agarose gels.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

What makes E-Gel agarose gels bufferless?

To create a bufferless system, each E-Gel cassette contains two unique ion exchange matrices that lie between the running gel and the electrodes. The ion exchange matrices provide a buffer-ion reservoir that supplies a continuous flow of Tris, acetate, and dye ions throughout the gel. This patented technology results in a sustained electric field with enhanced buffering capacity. E-Gel gels thus eliminate the need for you to prepare and dispose of liquid buffer, thus saving time and waste.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Gel Electrophoresis and Blotting Support Center.

What buffer should I use for my E-Gel agarose gel electrophoresis experiments?

The E-Gel agarose gel system is a precast bufferless TAE system that uses ion exchange matrices. The gels themselves are enclosed within a semi-UV-transparent cassette. This patented technology results in a sustained electric field with enhanced buffering capacity. E-Gel gels thus eliminate the need for you to prepare and dispose of liquid buffer, thus saving time and waste.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

What does intact RNA look like when run on an agarose gel?

Intact RNA should have a 2:1 ratio of 28S:18S bands. You may see a smear of RNA that extends from <9 kb to 0.5 kb, indicating the presence of mRNA in the sample. To see an image or to read more about RNA assessment, visit this website (https://www.thermofisher.com/us/en/home/references/protocols/nucleic-acid-purification-and-analysis/rna-protocol/agarose-gel-electrophoresis-of-rna.html).

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

How can I perform RNA electrophoresis?

For nondenaturing RNA electrophoresis, we recommend using our E-Gel Precast Agarose Gels. Please note that E-Gel Agarose Gels are not validated to be RNAse-free. However, many of our customers routinely use E-Gel Agarose Gels for RNA analysis with success. If RNA is run on an E-Gel Agarose Gel, any loading buffer that would be used for nondenaturing RNA electrophoresis should be fine.

For denaturing RNA electrophoresis, there are several denaturing agents to choose from, including formaldehyde, glyoxal, formamide, and methyl mercury. Denaturing conditions disrupt hydrogen bonding so that RNA runs without secondary structure, as single-stranded molecules.

For denaturing RNA electrophoresis, our E-Gel EX Agarose Gels can be used. The only denaturing agent that is compatible with the E-Gel EX system is formamide, 50-90%. Using other denaturing agents will result in poor band separation and morphology. Please note that we do not recommend running samples prepared in RNA loading buffer on the same gel with samples prepared in water. Please see below for the RNA loading buffer recipe and denaturing electrophoresis conditions:

RNA Loading Buffer:
Deionized formamide: 200 µL
10X MOPS-EDTA-Sodium Acetate Buffer (0.4 M MOPS, pH 7.0, 0.1 M sodium acetate, 10 mM EDTA): 40 µL
Deionized formaldehyde: 76 µL
Water: 14 µL

Denaturing Electrophoresis Conditions:
1. Mix 15 µL of RNA loading buffer with 1-5 µL of RNA (1-5 µg).
2. Heat samples at 65 degrees C for 10 min to denature RNA.
3. Place samples on ice immediately after heating.
4. Load entire sample onto an E-Gel EX agarose gel.
5. Electrophorese for 30 minutes.

For denaturing RNA electrophoresis under formaldehyde-free conditions, we recommend using our NorthernMax-Gly Kit (Cat. No. AM1946). With this kit, RNA samples are denatured in glyoxal/DMSO loading buffer and run on a glyoxal-containing agarose gel.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

Should I use agarose or polyacrylamide gels for DNA electrophoresis?

Agarose is commonly used as it is nontoxic, easy to use, and offers a broad range of separation. We offer precast E-Gel Agarose Gels or reagents to pour your own agarose gels. Polyacrylamide gels are typically used for high resolution of DNA molecules that range in size from 10-3,000 bp. We offer precast Invitrogen TBE polyacrylamide gels and UltraPure reagents.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

What is the function of the E-Editor 2.0 software?

The E-Editor 2.0 software allows you to quickly reconfigure digital images of E-Gel 48, E-Gel 96, E-PAGE 48 and E-PAGE 96 gel results for analysis and documentation. You can capture an image of the gel and then use the E-Editor 2.0 software to:
-Align and arrange the lanes in the image to any 48, 96, or 384 image
-Save the reconfigured image for further analysis
-Copy and paste selected lanes or the entire image into other applications for printing, saving, e-mailing, and/or publishing on the Web.

The E-Editor 2.0 does not take perform densitometry analysis from your gel images. The E-Editor 2.0 can be downloaded for free.

Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.

Do I need to pre-run my E-Gel agarose gels?

Pre-run is no longer necessary for any of our E-Gel agarose gels.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

Can E-Gel agarose gels be used with RNA samples?

E-Gel agarose gels are routinely used in-house in R&D and manufacturing for RNA analysis with excellent results. However, the gels are not guaranteed to be “RNAse-free”. The manufacturing process is designed to avoid contamination of any type, but not RNAses specifically.

If you do want to try it, any loading buffer that would be used for non-denaturing RNA electrophoresis should be fine for E-Gel agarose gels. Depending on your application, these gels may or may not be suitable for use.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

Can I run E-Gel agarose gels longer than recommended

Do not run E-Gel agarose gels longer than 40 min for the single comb gels or longer than 20 min for the double comb gels as longer run times will cause ions to get depleted and will damage the gel. Do not run E-Gel 48 agarose gels longer than 30 min or E-Gel 96 agarose gels longer than 20 min.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

What is the shelf-life of an E-Gel agarose gel?

All E-Gel gels are labeled with expiration dates.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

My E-Gel agarose gel is not running evenly. I am getting poor resolution or smearing of bands. What might be the problem?

Potential problems include:
- Loading too much DNA. Do not load more than the recommended amount.

- Samples with a high salt concentration. Samples containing > 50 mM NaCl, 100 mM KCl, 10 mM acetate ions, or 10 mM EDTA will cause loss of resolution.

- Samples may have been diluted in TAE instead of TE or water.

- Samples may have been pre-run; pre-running is not recommended for any of our E-Gel agarose gels.

- Sample was not properly loaded or had a very low volume of sample loaded. Load the recommended sample volume based on the gel type and loading method. For proper band separation, we recommend keeping sample volumes uniform.

- Bubbles may have been introduced while loading the samples. Bubbles will cause band distortion. Load deionized water or TE into any empty wells, and avoid introducing bubbles.

- A longer electrophoresis run time was used. Do not run the gel longer than the recommended time for each gel type. Longer run times cause an increase in the current, resulting in poor band migration or a melted gel. We recommend that you run single comb gels for 25-30 min (double comb gels 15-20 min) for straighter patterns. Do not run single comb gels longer than 40 minutes (20 minutes for double comb), or the gel will be damaged and resolution will be poor.

- Voltage or current too high. This should not be an issue with the E-Gel PowerBase power supply, which is pre-set with the proper conditions. However, researchers using the old E-Gel Base (the one that plugged into a separate power supply) should ensure that they run the gel at 60 to 70 volts (constant voltage) or 40-50 mA (constant current). Do not allow the current to exceed 60 mA.

- For E-Gel 96 Agarose Gels being run on an E-Base device, be sure you are running on the “EG” program rather than the “EP” program designed for E-PAGE gels.

- The gel was not electrophoresed immediately after sample loading (for best results, run gel within 1 min of loading).

- The gel may have been used beyond its expiration date. Check the expiration date.

Find additional tips, troubleshooting help, and resources within ourNucleic Acid Gel Electrophoresis and Blotting Support Center.

What grade of agarose is used in the E-Gel agarose gels?

The agarose used in the 0.8%, 1.2% and 2% E-Gel agarose gels is molecular biology grade, with a normal melting point and less than 0.25% ash content. For the 4% E-Gel agarose gels, we use a special high-resolution agarose.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

What buffer is used in the E-Gel agarose gel system?

The E-Gel agarose gel system uses TAE buffer.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

What are the dimensions of a single comb 12 well E-Gel agarose gel?

The dimensions of a single comb 12 well E-Gel agarose gel are as follows: The cassette itself is 8 cm by 10 cm and 0.6 cm thick. The workable gel area is 7.5 cm by 5.8 cm (from well to bottom). The thickness of the gel is estimated to be 0.4 cm. There are 12 wells and each well is 4 mm wide. The space between the wells is 1 mm.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

What ladder do you recommend for running RNA on Invitrogen E-Gel EX Agarose Gels?

Depending on the RNA fragment size, we recommend the following ladders:

• RiboRuler Low Range RNA ladder, (100 bp-1,000 bp) (Cat. No. SM1833)


• RiboRuler High Range RNA ladder, (200 bp-6,000 bp) (Cat. No. SM1823)

Can you provide me with the protocol to run E-Gel EX gels for RNA samples on the E-Gel PowerSnap Electrophoresis System?

Yes, the protocol for running E-Gel EX gels for RNA samples on the E-Gel PowerSnap Electrophoresis System can be found in the E-Gel Power Snap Electrophoresis System User Guide, on page 46 under Appendix E.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Gel Electrophoresis and Blotting Support Center.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Gel Electrophoresis and Blotting Support Center.

Can E-Gel EX Agarose Gels, 2% be used in a standard gel electrophoresis apparatus?

No. E-Gel EX Agarose Gels, 2% are only compatible with the E-Gel Power Snap Electrophoresis System (Cat. No. G8300).

What are the size dimensions of E-Gel EX Agarose Gels, 2% (Cat. No. G402022)?

Please see page 22 of the E-Gel Technical Guide for size dimensions (https://tools.thermofisher.com/content/sfs/manuals/egelguide_man.pdf).

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

Can I use the E-Gel Power Snap Electrophoresis Device (Cat. No. G8100) to image my own pour-your-own gels?

We do not recommend using the E-Gel Power Snap Electrophoresis Device to image pour-your-own gels.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

Can I use the E-Gel Power Snap Electrophoresis Device (Cat. No. G8100) to run my own pour-your-own gels?

We do not recommend using the E-Gel Power Snap Electrophoresis Device to run pour-your-own gels.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

Is E-Gel software compatible with Windows 10?

Yes, both the E-Gel GelCapture Acquisition Software and E-Gel GelQuant Express Analysis Software applications are compatible with Windows 10; however, the E-Gel GelQuant Express Analysis Software will require the purchase of an E-Gel Imager Quantitation USB dongle (Cat. No. 4466610):
https://www.thermofisher.com/order/catalog/product/4466610

Please visit the following link (https://www.thermofisher.com/us/en/home/life-science/dna-rna-purification-analysis/nucleic-acid-gel-electrophoresis/e-gel-electrophoresis-system/e-gel-imager-system/e-gel-software.html) and follow the instructions to download the software.
Note: Please make sure that you download the correct version of the E-Gel GelCapture Software based on the serial number of your instrument.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

How can I adjust the voltage on the E-Gel Power Snap Electrophoresis Device?

Each program on the E-Gel Power Snap Electrophoresis Device is optimized to obtain the best possible results. The device does not offer user-adjustable voltage functionality.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

I have an E-Gel Simple Runner Electrophoresis Device in my lab, are the new 11-well and 22-well E-Gel agarose gels compatible with it?

Due to the differences in dimensions, the new E-Gel 11-well and 22-well agarose gel formats are not compatible with the E-Gel Simple Runner Electrophoresis Device.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

Which DNA ladders do you recommend using with E-Gel EX agarose gels and E-Gel agarose gels with SYBR Safe stain?

To select the DNA ladder that yields the best resolution for your specific E-Gel, please refer to the table on page 45 of the E-Gel Power Snap Electrophoresis System user guide (https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0017050_egel_powersnapsystem_UG.pdf).

Can I use double comb E-Gel EX agarose gels or E-Gel agarose gels with SYBR Safe stain for RNA analysis?

E-Gel EX Double Comb agarose gels and E-Gel Double Comb agarose gels with SYBR Safe stain have not been tested for use in RNA applications. We recommend using E-Gel EX Single Comb agarose gels with 11 wells for RNA analysis.

I can't see any of my bands on my E-Gel agarose gel when I turn on the blue light. What should I do?

Follow recommended DNA dilutions and leave the gel to cool down on the bench or for a few minutes in the fridge. Please check troubleshooting tips provided in the manual.

Can I dispose of E-Gel EX agarose gels the same way as E-Gel agarose gels with SYBR Safe DNA Gel Stain?

No. Standard safety and hazardous waste disposal procedures should be followed when handling E-Gel EX agarose gels.

What is the difference between E-Gel agarose gels with SYBR Safe DNA Gel Stain and E-Gel EX agarose gels?

E-Gel EX agarose gels separate DNA faster, offer enhanced sensitivity, and provide added flexibility. The stain within the E-Gel EX agarose gels is SYBR Gold II which has similar spectral properties but increased sensitivity compared to SYBR Safe DNA Gel Stain. E-Gel EX agarose gels are especially suited for applications where high sensitivity is critical.

Can I run E-Gel agarose gels more than once?

No. E-Gel agarose gels have enough buffering capacity for one run only. Performance will be impaired with multiple runs.